An efficient synthesis method targeted to marine alkaloids marinacarbolines A-D and their antitumor activities

Article abstract of DOI:10.1080/10286020.2014.1003049

Marinacarbolines A-D are a series of marine β-carboline alkaloids isolated from actinomycete Marinactinospora thermotolerans of the deep South China Sea with antiplasmodial activities. In inhibition assays of in vitro growth of Plasmodium falciparum, marinacarbolines exhibited antiplasmodial activity against drug-sensitive line 3D7 and drug-resistant line Dd2 of P. falciparum. However, approaches for the synthesis of such useful compounds are very limited. In this work, we reported a simple, efficient, and versatile process to synthesize marinacarbolines A-D (1-4). On the basis of that, the antitumor activities of marinacarbolines in a structure-dependent manner were allowed to be unveiled.

Full text of DOI:10.1080/10286020.2014.1003049

This article was downloaded by: [University of Waterloo]
On: 31 March 2015, At: 19:40
Publisher: Taylor & Francis
Informa Ltd Registered in England and Wales Registered Number: 1072954 Registered
office: Mortimer House, 37-41 Mortimer Street, London W1T 3JH, UK
Journal of Asian Natural Products
Research
Publication details, including instructions for authors and
subscription information:
http://www.tandfonline.com/loi/ganp20
An efficient synthesis method targeted
to marine alkaloids marinacarbolines
A–D and their antitumor activities
Jun Lia, Yang Tanga, Hui-Juan Jina, Yi-Di Cuia, Li-Juan Zhanga &
Tao Jianga
a Key Laboratory of Marine Drugs, Chinese Ministry of Education,
School of Medicine and Pharmacy, Ocean University of China,
Qingdao 266003, China
Published online: 28 Jan 2015.
Click for updates
To cite this article: Jun Li, Yang Tang, Hui-Juan Jin, Yi-Di Cui, Li-Juan Zhang & Tao Jiang
(2015) An efficient synthesis method targeted to marine alkaloids marinacarbolines A–D and
their antitumor activities, Journal of Asian Natural Products Research, 17:3, 299-305, DOI:
10.1080/10286020.2014.1003049
To link to this article: http://dx.doi.org/10.1080/10286020.2014.1003049
PLEASE SCROLL DOWN FOR ARTICLE
Taylor & Francis makes every effort to ensure the accuracy of all the information (the
“Content”) contained in the publications on our platform. However, Taylor & Francis,
our agents, and our licensors make no representations or warranties whatsoever as to
the accuracy, completeness, or suitability for any purpose of the Content. Any opinions
and views expressed in this publication are the opinions and views of the authors,
and are not the views of or endorsed by Taylor & Francis. The accuracy of the Content
should not be relied upon and should be independently verified with primary sources
of information. Taylor and Francis shall not be liable for any losses, actions, claims,
proceedings, demands, costs, expenses, damages, and other liabilities whatsoever or
howsoever caused arising directly or indirectly in connection with, in relation to or arising
out of the use of the Content.
This article may be used for research, teaching, and private study purposes. Any
substantial or systematic reproduction, redistribution, reselling, loan, sub-licensing,
systematic supply, or distribution in any form to anyone is expressly forbidden. Terms &

Conditions of access and use can be found at http://www.tandfonline.com/page/terms-
and-conditions

Journal of Asian Natural Products Research, 2015
Vol. 17, No. 3, 299–305, http://dx.doi.org/10.1080/10286020.2014.1003049
An efficient synthesis method targeted to marine alkaloids
marinacarbolines A–D and their antitumor activities
Jun Li1, Yang Tang1, Hui-Juan Jin, Yi-Di Cui, Li-Juan Zhang* and Tao Jiang*
Key Laboratory of Marine Drugs, Chinese Ministry of Education, School of Medicine and
Pharmacy, Ocean University of China, Qingdao 266003, China
(Received 27 October 2014; final version received 7 December 2014)
Marinacarbolines A–D are a series of marine b-carboline alkaloids isolated from
actinomycete Marinactinospora thermotolerans of the deep South China Sea with
antiplasmodial activities. In inhibition assays of in vitro growth of Plasmodium
falciparum, marinacarbolines exhibited antiplasmodial activity against drug-sensitive
line 3D7 and drug-resistant line Dd2 of P. falciparum. However, approaches for the
synthesis of such useful compounds are very limited. In this work, we reported a simple,
efficient, and versatile process to synthesize marinacarbolines A–D (1–4). On the basis
of that, the antitumor activities of marinacarbolines in a structure-dependent manner
were allowed to be unveiled.
Keywords: marinacarbolines; marine b-carboline alkaloid; synthesis; antitumor
activities
1. Introduction
Very recently, Hibino and co-workers
have reported the first chemical synthesis
of marinacarbolines A–D from methyl
1-chloro-b-carboline-3-carboxylate in
four steps [5]. However, the required
compound methyl 1-chloro-b-carboline-
3-carboxylate was prepared in another six-
step sequence which is not an atom
economical process [5]. Our previous
studies on the structure–activity relation-
ship indicated that the electron-withdraw-
ing substitution at C-3 position of the b-
carboline alkaloids confers antitumor
activities [6]. Because of the limited
amount of natural occurring compounds,
the potential antitumor activities of mar-
inacarbolines A–D remain unknown.
Indole alkaloids are considered to be one
of the most important structural units
having great significance in medicinal
chemistry. The b-carboline alkaloids,
containing a tricyclic pyrido[3,4-b]indole
ring [1], displayed a wide range of
biological properties, such as antitumor,
anti-HIV, antimalarial [2], and neuropro-
tecting activities [3]. Because of their
practical importance, the b-carboline
alkaloids have drawn tremendous interest
from synthetic and medicinal chemists [1].
Marinacarbolines A–D (Figure 1, 1–
4), four new marine b-carboline alkaloids,
have been isolated from Marinactinospora
thermotolerans SCSIO 00652 obtained
from the deep South China Sea, and
exhibited antiplasmodial activities against
Plasmodium falciparum of both the drug-
sensitive (3D7) and drug-resistant (Dd2)
strains [4]. In 2013, Ju et al. [3]
successfully synthesized marinacarbolines
A–D through an enzymatic approach.
Given our longstanding interest in the
b-carboline alkaloids [7] and the intri-
guing biological profile of marinacarbo-
lines A–D, we undertook the development
of a scalable total synthesis of marinacar-
bolines A–D to provide sufficient quan-
tities for additional biological evaluation.
*Corresponding authors. Emails: lijuanzhang@ouc.edu.cn; jiangtao@ouc.edu.cn
q 2015 Taylor & Francis

300
J. Li et al.
R
O
O
NH
N
H
N
H
N
N
N
H
N
H
O
O
1 Marinacarboline A
2 Marinacarboline B
3 Marinacarboline C
R = OCH3
R = OH
R = H
4 Marinacarboline D
O
OH
N
N
H
O
5
Figure 1. The structures of b-carbolines derivatives (1–5).
We herein report the total chemical
synthesis of marinacarbolins A–D through
the Pictet–Splengler reaction from L-
tryptophan. In addition, antitumor activi-
ties of marinacarbolines A–D and com-
pound 5 against a human colon cancer cell
line (HCT116) and two human lung cancer
cell lines (H1299 and A549) were
investigated.
However, highly toxic iodine was
employed in the reaction process. The
modification method is to use p-toluene-
sulfonic acid as the acid catalyst to make
the cyclization and aromatization in one
step to form b-carboline directly with the
reaction between L-tryptophan and differ-
ent appropriate aldehydes [6]. This strat-
egy for the construction of the b-carboline
derivatives is efficient and straightforward,
and based on the principle, we previously
accomplished the total syntheses of marine
alkaloid pityriacitry [6] and eudistomins
Y1–Y7 [7].
2. Results and discussion
2.1. Chemistry
The traditional methods for the synthesis
of b-carboline alkaloids are mainly
focused on the two-step methods of
Pictet–Spengler and Bischler–Napier-
alski condensations [8–11], which is the
cyclization of L-tryptophan with different
appropriate aldehyde to provide a corre-
sponding diastereoisomeric mixture
of 1,2,3,4-tetrahydro-b-carboline-3-car-
boxylic acid as the first step, followed by
the aromatization of dehydrogenation to
yield b-carboline. Very recently, Ahmed
et al. [12] reported unexplored multi-
coupled domino reactions for the total
synthesis of marinacarboline and analogs.
The amide-substituted group at 3-
position of b-carboline is unique to marine
alkaloid marinacarbolines A–D. There-
fore, the b-carboline-3-carboxylic acid is
the key intermediate to facilitate the
chemical synthesis of marinacarbolines
A–D. Based on the established method,
b-carboline-3-carboxylic acid was formed
in lower yield as a by-product because the
carboxyl group at 3-position was easily
removed at the aromatization step [6].
Not surprisingly, our synthetic strategy for
the key intermediate of b-carboline-3-
carboxylic acid was envisioned to exploit

Journal of Asian Natural Products Research
301
O
COOH
NH2
COOCH3
O
a
b
NH2
N
N
H
N
H
N
H
O
6
7
8
R
O
c
N
1 Marinacarboline A R=OCH3
3 Marinacarboline C R=H
H
N
N
H
O
Scheme 1. Synthesis of marinacarbolines A (1) and C (3). Reagents and conditions: (a) HCl/
CH3OH, rt, 89%; (b) methyl–glyoxal/p-TSA/CH3OH, 648C, 120 min, 90%; (c) triethylamine/4-
methoxyphenethylamine/CHCl3, 508C, for 1, 85%; triethylamine/2-phenylethylamine/CHCl3, 508C,
for 3, 82%.
the lessons learned in our earlier synthesis,
with important modifications to ensure
efficient access to the natural product
(Scheme 1).
of 1-acetyl-3-chlorocarbonyl-b-carboline
with the corresponding amine (Scheme 2).
At 608C, 1-acetyl-3-(methoxycar-
bonyl)-b-carboline (8) was treated with
18% aqueous NaOH to remove the methyl
group and to form 1-acetyl-3-carboxy-b-
carboline (8a) in a yield of 92%. 8a was then
treated with oxalyl chloride in dichloro-
methane to obtain 1-acetyl-3-chlorocarbo-
nyl-b-carboline (8b). Finally, 8b was used
to react with tyramine or tryptamine in the
presence of pyridine and CH2Cl2 as solvent
to afford a target compound marinacarbo-
lines B or D (2 or 4). The reaction routes are
outlined in Scheme 2.
To obtain 1-acetyl-3-carboxy-b-carbo-
line, L-tryptophan methyl ester (7) was
used to react with methyl-glyoxal in
methanol, and p-toluenesulfonic acid was
used as the catalyst for 2 h at 648C.
By using this condition, 1-acetyl-3-(meth-
oxycarbonyl)-b-carboline (8) was formed
in one-pot in a yield of 90%.
Subsequently, marinacarbolines A and
C (1 and 3) were synthesized by the
reaction of 1-acetyl-3-(methoxycarbonyl)-
b-carboline (8) with different appropriate
amines, 2-phenylethylamine and 4-meth-
oxyphenethylamine, in CHCl3 at 508C,
respectively. The reaction routes are out-
lined in Scheme 1.
2.2. Biological activities
We measured the percentage of viable
cancer cells after 48 h exposure to the
b-carbolines derivatives, marinacarbolines
A–D (1–4), and the key intermediate 5 at
a concentration of 50 mM as compared
with the compound-free control that is
100% viability (Figure 2) in the three
cancer cell lines including two lung cancer
(A549 and H1299) and one colon cancer
(HCT116) cells. Compounds 4 and 5
showed considerable cytotoxicities in the
three cancer cell lines tested.
However, marinacarbolines B and D (2
and 4) could not be obtained from 1-
acetyl-3-(methoxycarbonyl)-b-carboline
(8), because tyramine and tryptamin in the
ester exchange reaction of amine were
easy to form by-product, and the reaction
activity was lower when heated up to
508C. To increase the reaction yield, the
target compounds, marinacarbolines B and
D, were synthesized through the reaction

302
J. Li et al.
COOCH3
COOH
COCl
a
b
N
N
N
N
N
N
H
H
H
O
O
O
8b
8a
8
OH
O
N
2 Marinacarboline B
4 Marinacarboline D
H
N
c
N
H
O
d
O
NH
N
H
N
N
H
O
Scheme 2. Synthesis of marinacarbolines B (2) and D (4). Reagents and conditions: (a) NaOH/
H2O/CH3OH, 608C, 92%; (b) C2Cl2O2/ CH2Cl2; (c) tyramine/pyridine/CH2Cl2; 68%; (d)
tryptamine/pyridine/DMF/CH2Cl2; 71%.
Subsequently, the IC50 in the three
cancer lines was explored further using
doxorubicin as a positive control (Table 1).
All the b-carbolines derivatives showed
moderate cytotoxicities in each tested cell
line. Interestingly, the IC50 value for each
compound was not proportional to the data
shown in Figure 1, this indicated that the
concentration-dependent cytotoxicity
curve was unique to each compound. The
data in Table 1 showed that compound 5
with carbonyl group and compound 4 with
amide of tryptamine at the 3-position of b-
carbolines were more toxic than other
compounds in H1299 and HCT116 cells
with IC50 which ranged from 11 to 18 mM
and less toxic than the positive control of
doxorubicin.
Figure 2. The percentage of viable cancer cells after 48 h exposure to compounds 1–5 at a
concentration of 50 mM compared with the compound-free control (100% viability). The compounds
were formulated initially in DMSO and then diluted in complete growth media (details are in
experimental protocols) (B, A549; , HCT116; A, H1299).

Journal of Asian Natural Products Research
303
Table 1. IC50 values of compounds 1–5 in cancer cells A549, H1299, and HCT116 after 48 h
exposure to the compounds.
IC50(mM)
Compound
A549
H1299
HCT116
Marinacarboline A (1)
Marinacarboline B (2)
Marinacarboline C (3)
Marinacarboline D (4)
intermediate (5)
33.9 ^ 7.95
57.6 ^ 1.50
47.5 ^ 1.50
29.9 ^ 6.13
32.9 ^ 2.60
0.32 ^ 0.06
26.9 ^ 4.75
63.7 ^ 3.13
47.5 ^ 3.14
15.0 ^ 0.15
14.7 ^ 0.25
6.70 ^ 0.59
24.4 ^ 2.00
37.9 ^ 0.86
37.4 ^ 2.23
23.9 ^ 3.81
18.7 ^ 2.17
0.20 ^ 0.09
Doxorubicin
3. Experimental
3.2. General procedure for 1-acetyl-3-
(methoxycarbonyl)-b-carboline synthesis
(8)
3.1. Materials and methods
Melting points were recorded on a micro
melting point apparatus MP-500D and are
uncorrected (Shanghai Instrument Physical
Optics Instrument Co. Ltd., Shanghai,
China). NMR spectra were recorded on a
JeolJNM-ECP spectrometerat600 MHz for
1H NMR and 150 MHz for 13C NMR with
TMS as the internal standard (JEOL Ltd.,
Tokyo, Japan). Chemical shifts are
expressed in d (ppm) and coupling constants
(J) in Hz. Multiplicity is indicated as
follows: s (singlet), d (doublet), t (triplet),
dd (doublet of doublets), brs (broad singlet),
and so on. Mass spectra were recorded using
a Q-TOF Ultimae Global by chemical
ionization (Thermo Fisher Scientific, Inc.,
Waltham, MA, USA). Column chromatog-
raphy was carried out using silica gel (200–
300 mesh, Qingdao Marine Chemical Fac-
tory, Qingdao, China). The starting
materials and reagents purchased from
commercial suppliers were used without
further purification. All reactions were
monitored by a thin-layer chromatography
(TLC), on aluminum sheets (Silica gel 60-
To a solution of L-tryptophan methyl ester
(0.18 g, 0.67mmol) in CH3OH (40 ml), p-
toluenesulfonic acid monohydrate (0.14 g,
0.67 mmol) and 40% methyl–glyoxal
(1.2 ml, 6.67 mmol) were added. The
solution was stirred at 648C for 2 h and the
completionofthereactionwasmonitoredby
TLC. The resulting suspended solution was
poured into 100 ml of ice water and the solid
materials were collected by filtration and
driedinvacuumtoaffordayellowsolid.The
crude products were purified by flash
column chromatography using silica gel as
the stationary phase and using ethyl acetate/
hexane (1:2) as the mobile phase to provide
compound 8.
1-Acetyl-3-(methoxycarbonyl)-b-car-
boline (8): yellow solid; yield 90%; Mp
1
224–2278C; H NMR (600 MHz, CDCl3)
d: 10.51 (s,1H), 9.02 (s, 1H), 8.20 (d,
J ¼ 7.7 Hz, 1H), 7.58–7.65 (m, 2H), 7.40
(t, J ¼ 7.7 Hz, 1H), 4.09 (s, 3H), 2.97 (s,
3H). ESI-MS: (m/z) 269 [M þ H]þ, 291
[M þ Na]þ.
F254, E. Merck, Darmstadt, Germany).
Compounds were visualized by UV light.
All reaction solvents were dried prior to use,
according to the standard procedures. All
primary reagents were commercially avail-
able. Silica gel chromatography solvents
were of analytical grade.
3.3. General procedure for
marinacarboline A and marinacarboline C
To the solution of 1-acetyl-3-(methoxy-
carbonyl)-b-carboline (1 g, 3.7 mmol) in
CHCl3 (30 ml), triethylamine (1.5 g,

304
J. Li et al.
3.7 mmol) and 2-phenylethylamine
(1.88 ml, 6.4 mmol) were added dropwise.
The reaction mixture was heated at 508C for
6 h and the completion of the reaction was
monitored by TLC. After cooling to room
temperature, the solvent was removed under
reduced pressure. The residue was poured
into water (100ml) and extracted with ethyl
acetate (20 ml £ 3). The combined organic
phase was washed with 5% aqueous
NaHCO3 (20 ml £ 3), H2O (20 ml £ 3),
and brine (20 ml £ 3), dried over anhydrous
MgSO4 and the solvent was removed under
reduced pressure to afford a yellow crude
product. The residue was purified by flash
column chromatography using silica gel as
the stationary phase and using ethyl acetate/
hexane (1:15) as the mobile phase to
provide compound 1.
Marinacarboline A (1): Yellow solid;
yield 85%; 1H NMR (600 MHz, CDCl3) d:
10.39 (s, 1H), 9.09 (s, 1H), 8.22 (d,
J ¼ 7.7 Hz, 1H), 8.07 (t, J ¼ 5.5 Hz, 1H),
7.59–7.64 (m, 2H), 7.38 (t, J ¼ 7.7 Hz,
1H), 7.23–7.26 (m, 2H), 6.90 (d,
J ¼ 8.8 Hz, 2H), 3.79–3.83 (m, 5H), 2.96
(t, J ¼ 6.6 Hz, 2H), 2.78 (s, 3H); 13C NMR
(150 MHz, CDCl3) d: 202.3, 164.6, 158.5,
141.6, 139.3, 136.3, 133.5, 132.7, 131.1,
130.0, 129.8, 122.4, 121.6, 121.1, 118.4,
114.3, 112.2, 55.4, 40.8, 35.1, 25.8.
CH3OH (10 ml), 18% aqueous NaOH
(5.5 ml) was slowly added. After the
addition, stirring was continued for 2 h at
608C. The completion of the reaction was
monitored by TLC. Subsequently, concen-
trated hydrochloric acid was slowly added
to the reaction solution to adjust the pH to 1,
and the resulting reaction mixture was
filtered and dried in vacuum to afford a
white crude product. The crude product
was purified by flash column chromatog-
raphy using silica gel as the stationary
phase and using ethyl acetate/methanol/
formic acid (50:2:1) as the mobile phase to
provide the 1-acetyl-3-carboxy-b-carbo-
line (8a) as a white solid in a yield of 92%.
To a solution of 8a (1.0g, 3.7 mmol) in
anhydrous CH2Cl2 was added (COCl)2
(1.1ml, 11.1mmol). The above mixture
was stirred at room temperature for about
1 h and the completion of the reaction was
monitored by TLC. The resulting solution
was evaporated under reduced pressure. The
residue was solved in anhydrous CH2Cl2
and tryptamine (62 mg, 0.39mmol) and
pyridine (94 ml, 1.17mmol) were added.
In addition, stirring was continued over-
night at room temperature. The mixture was
poured into 10 ml of ice water and
extracted with ethyl acetate (30 ml £ 2).
The combined organic phase was washed
with 2 N aqueous hydrochloric acid (20 ml
£ 3), H2O (20 ml £ 3), and brine (20 ml
£ 3), dried over anhydrous MgSO4 and the
solvent was removed under reduced
pressure to afford a yellow crude product.
The crude product was purified by flash
column chromatography using silica gel as
the stationary phase and ethyl acetate/
hexane (1:10) as the mobile phase to
provide the title compound 4.
Marinacarboline C (3): Yellow solid;
yield 82%; 1H NMR (600 MHz, CDCl3) d:
10.38 (s, 1H), 9.08 (s, 1H), 8.21 (d,
J ¼ 7.7 Hz, 1H), 8.07 (t, J ¼ 5.5 Hz, 1H),
7.58–7.66 (m, 2H), 7.32–7.40 (m, 5H),
7.26–7.29 (m, 1H), 3.86 (t, J ¼ 7.2 Hz,
2H), 3.02 (t, J ¼ 7.1 Hz, 2H), 2.75 (s, 3H);
13C NMR (150 MHz, CDCl3) d: 202.4,
164.7, 141.7, 139.2, 133.6, 136.4, 132.7,
131.0, 129.8, 129.1, 129.0, 128.9, 128.8,
126.7, 122.4, 122.4, 121.2, 118.4, 112.3,
40.6, 36.0, 25.8.
Marinacarboline D (4): Yellow solid;
1
yield 68%; H NMR (600 MHz, DMSO-
d6) d: 12.17 (s, 1H), 10.90 (s, 1H), 9.11 (s,
1H), 8.78 (t, J ¼ 6.1 Hz, 1H), 8.44 (d,
J ¼ 8.3 Hz, 1H), 7.83 (d, J ¼ 6.1 Hz, 1H),
7.67 (d, J ¼ 6.1 Hz, 1H), 7.62 (t,
J ¼ 7.7 Hz, 1H), 7.36 (d, J ¼ 8.1 Hz,
1H), 7.35 (t, J ¼ 7.5 Hz, 1H), 7.27
3.4. General procedure for
marinacarboline D and marinacarboline B
To the solution of 1-acetyl-3-(methoxycar-
bonyl)-b-carboline (8, 0.2 g, 0.75 mmol) in

Journal of Asian Natural Products Research
305
(s, 1H), 7.08 (t, J ¼ 7.7 Hz, 1H), 6.98–
7.01 (m, 1H), 3.72 (q, J ¼ 6.6 Hz, 2H),
3.06 (t, J ¼ 7.7 Hz, 2H), 2.83 (s, 3H); 13C
NMR (151 MHz, CDCl3) d: 201.1, 164.1,
142.4, 138.8, 136.5, 134.9, 134.0, 132.0,
129.3, 127.4, 122.9, 122.3, 121.1, 120.9,
120.4, 118.6, 118.4, 118.0, 113.4, 111.8,
111.5, 39.6, 26.1, 25.4.
other b-carbolines possessing an amino
acid side chain, but also be effective in the
synthesis of other complex natural pro-
ducts. The chemically synthesized mar-
inacarbolines A–D and other analogs
made wider bioassay studies possible.
Funding
Marinacarboline B (2): Yellow solid;
yield 72%; 1H NMR (600 MHz, DMSO-d6)
d:12.17 (s, 1H) 9.23 (s, 1H), 9.08 (s, 1H),
8.68 (t, J ¼ 5.9 Hz, 1H), 8.45 (d,
J ¼ 8.1 Hz, 1H), 7.82 (d, J ¼ 8.2 Hz, 1H),
7.61 (t, J ¼ 8.2 Hz, 1H), 7.32–7.35 (m,
2H), 7.11 (d, J ¼ 8.3 Hz, 2H), 6.63 (d,
J ¼ 8.3 Hz, 2H), 3.58 (q, J ¼ 6.6 Hz, 2H),
2.87 (s, 3H), 2.80 (t, J ¼ 7.4 Hz, 2H); 13C
NMR (150 MHz, CDCl3) d: 203.3, 166.1,
141.4, 137.1, 136.5, 136.5, 135.3, 131.9,
129.8, 129.7, 122.1, 122.6, 121.7, 121.6,
121.3, 121.2, 120.9, 120.9, 112.4, 52.8,
29.9, 25.7.
This research was supported by Natural Science
Foundation of China [grant number 21171154],
[grant number 91129706]; and NSFC-Shan-
dong Joint Fund for Marine Science Center
[U1406402].
Note
1. These authors contributed equally to this
work.
References
[1] R.H. Cao, W.L. Peng, Z.H. Wang, and
A.L. Xu, Curr. Med. Chem. 14, 479(2007).
[2] J.D. Panarese and S.P. Waters, Org. Lett.
12, 4086 (2010).
[3] Q. Chen, C.T. Ji, Y.X. Song, H.B. Huang,
J.Y. Ma, X.P. Tian, and J.H. Ju, Angew.
Chem. Int. Edit. 52, 9980 (2013).
[4] H.B. Huang, Y.L. Yao, Z.X. He,
T.T. Yang, J.Y. Ma, X.P. Tian, Y.Y. Li,
C.G. Huang, X.P. Chen, W.J. Li, S. Zhang,
C.S. Zhang, and J.H. Ju, J. Nat. Prod. 74,
2122 (2011).
[5] S. Tagawa, T. Choshi, A. Okamoto,
T. Nishiyama, S. Watanabe, N. Hatae, and
S. Hibino, Heterocycles 87, 357 (2013).
[6] P.Y. Zhang, X.F. Sun, B. Xu, K. Bijian,
S.B. Wan, G.G. Li, M. Alaoui-Jamali, and
T.JiangEur,J.Med.Chem.46, 6089(2011).
[7] H. Jin, P. Zhang, K. Bijian, S. Ren,
S. Wan, M.A. Alaoui-Jamali, and
T. Jiang, Mar. Drugs 11, 1427 (2013).
[8] E.D. Cox and J.M. Cook, Chem. Rev. 95,
1797 (1995).
3.5. Biological activities
A549 (2000 cells/well), H1299 (2000 cells/
well), and HCT116 (2000 cells/well) cells
were seeded in 96-well plates. After 24 h,
cells were treated with serial concen-
trations of b-carbolines derivatives in
200 ml of serum complete media. After
48 h of treatment, 20 ml of resazurin (2 mg/
ml dissolved in water, Sigma, St Louis,
MO, USA) cells was added to the media.
After 16 h incubation at 378C, the fluor-
escent signal was monitored using 544 nm
excitation wavelength and 595 nm emis-
sion wavelength by a Spectramax M5 plate
reader (Molecular Devices, Sunnyvail, CA,
USA). The relative fluorescence unit
generated from the assay was proportional
to the number of living cells in each well.
[9] R. Faust, P.J. Garratt, R. Jones, L.K. Yeh,
A. Tsotinis, M. Panoussopoulou, T. -
Calogeropoulou, and M.T. Teh, J. Med.
Chem. 43, 1050 (2000).
[10] I. Ralonilic, M. Castillo, and J.T. Lopez,
J. Heterocycl. Chem. 17, 595 (1980).
[11] I. Nemet and L. Varga-Defterdarovic,
Bioorg. Med. Chem. 16, 4551 (2008).
[12] N. Battini, A.K. Padala, N. Mupparapu,
R.A. Vishwakarma, and Q.N. Ahmed,
RSC Adv. 4, 26258 (2014).
4. Conclusions
In summary, an efficient synthetic method
for marinacarbolines A–D was developed.
It was expected that this methodology
could not only be used in the synthesis of